Our goal is to develop a better understanding of cellular mechanisms that regulate aqueous humor secretion and an improved understanding of Na,K-ATPase function. The application focuses on the idea that function of Na, K-ATPase and other transport proteins can be modulated by changes in cytoplasmic pH (pHi). In recent studies we discovered that carbonic anhydrase inhibitors (CAIs) lower phi in cultured nonpigrnented ciliary epithelial (NPE) cells. In Aim I we propose studies to examine how cytoplasmic acidification alters Na, K-ATPase function. Experiments will be conducted to determine whether cytoplasmic acidification inhibits Na, K-ATPase activity via a tyrosine kinase mechanism. Studies will also be done to determine whether the pattern of changes in aqueous humor composition in rabbits treated with CAIs is consistent with altered Na, K-ATPase function in ciliary epithelium. Preliminary Studies also show sodium orthovanadate reduces pHi in NPE and experiments will be carried out to explore whether the pH change could be linked to the ability of sodium orthovanadate to reduce lOP. Since we propose changes of pHi could change the activity of Na, K-ATPase and other transport proteins, we consider H+-ATPase, a proton export pump, to be important to NPE function. Indeed lOP is reduced by the selective H+-ATPase inhibitor bafilomycin A1. Results from recent studies with angiotensin II and calcium channel blockers indicate the existence of cellular mechanisms that control H+-ATPase. In Aim II we propose experiments to learn more about Hv-ATPase and its control mechanisms. Studies will be conducted to examine the effects of cAMP and protein kinase A activation on Hv-ATPase and also to determine whether nitric oxide and vanadate cause inhibition of H+-ATPase. In summary, we propose to examine how ion transport mechanisms in the NPE are set up to respond to cytoplasmic pH changes and to study how pHi is controlled in the NPE. Improved basic science knowledge in this area might lead to the future development of lOP-lowering drugs that work by interfering with cell pH balance.